Part:BBa_K3402057

Double-gene editing cassette

This device is composed of 50bp-upPXA1(BBa_K3402037), hph(BBa_K3402012), 50bp-doPXA1(BBa_K3402038), Ptef1(BBa_K3402007), sgPXA1(BBa_K3402039), sgGFP(BBa_K3402024), Tsyn7(BBa_K3402001), upGFP(BBa_K3402025), doGFP(BBa_K3402026).

Usage and Biology

We knock out the PXA1 and GFP gene by this device. In the meantime, we insert hph gene at the origin PXA1 site by homology directed repair, which is to use hph as the marker gene to know if it is successful to knock PXA1.
At last, we observe the strains that growed on the plate coated with hygromycin and check if strains can also lose the function of expressing green fluorescence. Then, we get the conclusion that the success of double gene editing was proved.
The double gene-editing efficiency was 99%.

Fig. a)Strains can grow on the plate coated with hygromycin. b)Most of strains also lose the function of expressing green fluorescence.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal XhoI site found at 2171
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI.rc site found at 2807